Methods of breast cancer detection

Abstract

Methods of diagnosing breast cancers are provided. The methods include analyzing the expression patterns of biomarkers in a plurality of cells to distinguish between invasive and non-invasive cancers, usual and atypical hyperplasias, and basal-like breast cancers. Breast cancers are diagnosed based upon different combinations of the biomarkers found in nipple aspirate fluid.

1 .- 22 . (canceled) 
     
     
         23 . A method of classifying a breast cancer, comprising:
 a) contacting a cell of a nipple aspirate fluid sample absorbed onto an absorbent paper with antibodies that bind to CK5, CK14, CK7, CK18, and p63, wherein the absorbent paper is sized to cover a nipple;   b) detecting binding of one or more of the antibodies to said cell; and   c) classifying the cancer based upon the binding pattern of the primary antibodies;   wherein the cell derived from the nipple aspirate fluid (NAF) sample is not a tissue.   
     
     
         24 . The method of  claim 23 , wherein the absorbent paper is from about 1.0 to about 3.0 inches in diameter and from about 0.01 to about 0.1 inches thick. 
     
     
         25 . The method of  claim 23 , wherein detecting binding of the one or more antibodies further comprises staining the cells with a stain selected from among horseradish peroxidase, alkaline phosphatase, diaminobenzidine, Fast Red, hematoxylin, eosin and a combination thereof. 
     
     
         26 . The method of  claim 23 , further comprising washing the absorbent paper and collecting the effluent. 
     
     
         27 . The method of  claim 23 , wherein the absorbent paper comprises microcellulose, mixed cellulose ester, or nitrocellulose. 
     
     
         28 . The method of  claim 23 , wherein the absorbent paper is a device as illustrated in  FIG. 1 . 
     
     
         29 . The method of  claim 23 , further comprising classifying the breast cancer as basal-like if an anti-CK5 antibody, an anti-CK14 antibody, and optionally an anti-p63, primary antibody binds to the cell. 
     
     
         30 . The method of  claim 23 , further comprising classifying the breast cancer as luminal if (i) an anti-CK7 antibody and an anti-CK18 primary antibody bind to the cell, and (ii) an anti-CK5 antibody, and anti-CK14 antibody, and an anti-p63 antibody do not bind to the cell. 
     
     
         31 . The method of  claim 23 , further comprising classifying the breast cancer as usual ductal hyperplasia if an anti-CK5 antibody, an anti-CK14 antibody, an anti-CK7 antibody, an anti-CK18 antibody, and an anti-p63 primary antibody bind to the cell. 
     
     
         32 . The method of  claim 23 , further comprising classifying the cancer as atypical ductal hyperplasia if (i) the anti-CK7 antibody and anti-CK18 antibody, and optionally the anti-p63 antibody, bind to the cell, and (ii) the anti-CK5 antibody and anti-CK14 antibody do not bind to the cell. 
     
     
         33 . The method of  claim 23 , further comprising classifying the cancer as invasive if (i) the sample comprises more than one cell and (i) the ratio of cells binding the anti-CK5 antibody, anti-CK14 antibody, and anti-p63 antibody to cells binding the anti-CK7 antibody and anti-CK18 antibody is less than or equal to an invasive control. 
     
     
         34 . The method of  claim 23 , further comprising classifying the cancer as non-invasive if: (i) the sample comprises more than one cell, and (ii) the ratio of cells binding the anti-CK5 antibody, anti-CK14 antibody, and anti-p63 antibody to cells binding the anti-CK7 antibody and anti-CK18 antibody is greater than or equal to a non-invasive control. 
     
     
         35 . (canceled) 
     
     
         36 . A composition comprising (a) at least one cell derived from nipple aspirate fluid absorbed onto an absorbent paper comprising microcellulose, mixed cellulose ester, or nitrocellulose for absorbing a nipple aspirate fluid sample, wherein the absorbent paper is sized to cover a nipple; and (b) antibodies that bind to an antigens on a cell in the nipple aspirate fluid sample, wherein the antigens are selected from: CK5, CK14, CK7, CK18, p63, CK7 and CK18. 
     
     
         37 . A system for classifying a breast cancer, comprising:
 a) an absorbent paper comprising microcellulose, mixed cellulose ester, or nitrocellulose for absorbing a nipple aspirate fluid sample, wherein the absorbent paper is sized to cover a nipple; and   b) antibodies that bind to an antigens on a cell in the nipple aspirate fluid sample, wherein the antigens are selected from: CK5, CK14, CK7, CK18, p63, CK7 and CK18.   
     
     
         38 . The system of  claim 37 , further comprising a light microscope or an automated system for visualizing the antibodies bound to a cell in the nipple aspirate fluid sample. 
     
     
         39 . The system of  claim 37 , wherein the absorbent paper is from about 1.0 to about 3.0 inches in diameter and from about 0.01 to about 0.1 inches thick. 
     
     
         40 . The system of  claim 37 , further comprising means for visualizing the antibodies bound to the cell in the nipple aspirate fluid sample. 
     
     
         41 . The system of  claim 40 , wherein the means for visualizing the antibodies bound to the cell in the nipple aspirate fluid sample is one or more stains. 
     
     
         42 . The system of  claim 41 , wherein the stain is selected from horseradish peroxidase, alkaline phosphatase, diaminobenzidine, Fast Red, hematoxylin, eosin or a combination thereof. 
     
     
         43 . (canceled) 
     
     
         44 . The system of  claim 37 , further comprising: an optionally networked computer processing device configured to perform executable instructions; and a computer program, the computer program comprising a software module executed by the computer processing device to apply a model or algorithm for analyzing said cells. 
     
     
         45 .- 52 . (canceled)